N乙酰基-L-半胱氨酸对苯乙烯致耳蜗毛细胞损伤的防护作用

L-NAC protect hair cells in the rat cochlea from injury of exposure to styrene

目的:观察N乙酰基-L-半胱氨酸(L-NAC)对苯乙烯致耳蜗毛细胞损伤的防护作用。方法:入选成年Long Evens大鼠17只,随机分为2组:胃饲苯乙烯的对照组8只和苯乙烯加L-NAC治疗的实验组9只。胃饲苯乙烯400mg/kg(2g苯乙烯溶于1ml橄榄油),每日1次,每周5d,共3周。胃饲苯乙烯后,实验组动物腹腔注射L-NAC(325mg/kg,生理盐水稀释),对照组腹腔注射等量生理盐水。胃饲苯乙烯、苯乙烯加L-NAC治疗前及3周后,应用电位反应测听仪检测短声诱发的动物双侧ABR阈值。大鼠听觉功能检测后,断头处死。3种荧光染料用于苯乙烯及苯乙烯加L-NAC治疗3周后的耳蜗基膜染色,DNA标记物PI、原位末端标记和异硫氰酸荧光素标记的鬼笔环肽染色分别用于观察毛细胞核和纤毛、表皮板形态学变化。荧光显微镜下自耳蜗顶回到底回依次计数损伤(核固缩、核肿胀和核缺失)的毛细胞。结果:实验组L-NAC治疗3周后,短声诱发的平均ABR阈移数值明显低于对照组(P<0.05)。荧光显微镜下定量观察发现,对照组外毛细胞核损伤(核固缩、核肿胀和核缺失)平均数目为28.3%,而实验组仅为10.6%(P<0.01)。实验组核固缩、核肿胀外毛细胞数目分别比对照组数目减少78%(P<0.01)和23%(P>0.05)。结论:本实验结果表明,L-NAC对苯乙烯致耳蜗毛细胞损伤具有较好的防护作用,其抗细胞凋亡作用更为明显。

Objective：To observe the effects of N-acetyl-L-cysteine（L-NAC） protect hair cells in the rat cochlea from injury of exposure to styrene.Method：Seventeen adult Long Evans rats were used in present study.The animals were randomly assigned into test group（n=9） and control group（n=8）.The animals were exposed to styrene by gavage at 400 mg/kg（2 g styrene was mixed with 1ml olive oil）.Test group animals received styrene exposure plus L-NAC 325 mg/kg（L-NAC was dissolved in physiological saline solution） by intraperitoneal injection.Treatment was performed once a day,5 days per week for 3 weeks.Control group animals received the same volume of saline injection on an identical time schedule used for the test group.The auditory brainstem response（ABR） thresholds of both ears elicited with clicks were measured before and at the end of the 3-week styrene or styrene plus L-NAC treatment.After hearing was re-assessed,animals were sacrificed and cochleae were quickly removed from the skull.Following fixation,whole specimens comprising the basilar membrane with Corti＇s organ were separated from the modiolus.The organs of Corti were stained with propidium iodide（PI） and the TUNEL assay to visualize the morphologic viability of hair cell nuclei,FITC-labeled phalloidin,a F-actin intercalating fluorescent probe used to visualize the morphologic viability of cuticular plate and the stereocilia in the hair cells.Each organ of Corti was thoroughly examined using fluorescence microscopy.The numbers of damaged OHCs（apoptotic,necrotic and missing OHCs） were documented.Result：There was a statistically significant decrease in ABR threshold shift（P0.05）in the styrene-plus-L-NAC treated animals.The average percentage of damaged OHCs in the stytene-treated animals was 28.3%.In contrast,the average percentage of OHC damage in the styrene-plus-L-NAC treatmed group was only 10.6%（P0.01）.The percentage of reduction in the number of apoptotic cells in styrene-plus-L-NAC treated group was 78%（P0.01）.However,the mean reduction of necrotic cells was only 23%（P0.05）.Conclusion：The results indicate that the treatment with L-NAC may effectively protect against the styrene-induced hair cells damage and preferably reduce the number of apoptotic OHCs.